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Direct Observation of Enzymes Replicating DNA Using a Single-molecule DNA Stretching Assay
Published on: March 24, 2010
Enzymes involved in DNA synthesis in the testes are regulated by temperature in vitro
M Fujisawa1, A Hayashi, H Okada
1Department of Urology, Kobe University School of Medicine, Japan.
Objective:
Temperature is considered to be an important local regulating factor of testicular function. We investigated whether a high temperature would affect the activity of DNA polymerases alpha, beta, and gamma (EC2.7.7.7) and DNA topoisomerase I in the testes which are required for germ cell differentiation.
Methods:
An in vitro rat testis tissue culture system was used to evaluate the activity of DNA polymerases alpha, beta, and gamma and DNA topoisomerase I after incubation at 31 and 37 degrees C. The expression of proliferating cell nuclear antigen was evaluated immunohistochemically in the testis.
Results:
Incubation at 37 degrees C significantly reduced the activity of all four enzymes as compared with incubation at 31 degrees C. Reducing the temperature to 31 degrees C after prior incubation at 37 degrees C partially restored DNA polymerase alpha activity. Expression of proliferating cell nuclear antigen demonstrated a reduction in DNA synthesis in the seminiferous tubules after incubation at 37 degrees C CONCLUSION: These findings indicate that a high temperature of 37 degrees C reduces the activity of the enzymes involved in the testicular synthesis of DNA which may cause the impairment of spermatogenesis.
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